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One of the T suppressor circuits induced by picrylsulphonic acid includes the T suppressor cell (Ts-eff) which acts at the efferent stage of the contact sensitivity reaction and produces antigen-specific T suppressor factor (TsF). This factor does not act directly but arms a T acceptor cell (Tacc). This Tacc liberates a non-specific inhibitor when it is armed with TsF and then exposed to picrylated cells sharing the I-J genotype of the source of the TsF. This paper investigates the role of I-J region gene products in this T suppressor circuit. Two approaches were used. Syngeneic CBA (H-2k) lymphocytes were separated into I-J+ and I-J- cells by treatment with anti-I-Jk serum followed by panning on anti-immunoglobulin plates. The cells were then picrylated and used as a source of antigen. Alternatively, B10.A congeneic mice syngeneic (SR) or allogeneic (3R) with CBA at the I-J locus were picrylated and used similarly. The main findings were as follows. (i) The intravenous injection of picrylated I-J+ spleen cells but not a similar number of I-J- cells induced Ts-eff which blocked the transfer of contact sensitivity. Picrylated unseparated cells syngeneic, but not allogeneic, at the I-J locus were also effective. (ii) It is known that the lymphocytes of mice injected with picrylsulphonic acid and then re-exposed to antigen by painting with picryl chloride liberate TsF in vitro. The re-exposure to antigen can be replaced by the intravenous injection of picrylated I-J+ cells or by cells syngeneic at the I-J locus the day before harvesting the spleen cells. (iii) The release of non-specific inhibitor by Tacc armed with TsF requires exposure to picrylated I-J+ cells or cells syngeneic at the I-J locus. The requirement for antigen on a cell bearing syngeneic I-J suggests that antigen together with I-J is an activation signal in this T-cell circuit. The simplest explanation is that the receptor of the pristine Ts and of the mature Ts-eff is similar to T suppressor factor. [ABSTRACT FROM AUTHOR]

Lymph node cells from immunized rabbits were cultured with varying concentrations of antigen in preheated (56°, 30 minutes) autologous serum which had been collected before immunization. [3H]Thymidine was present for the last 6 hours of the 24-hour culture period and the radioactive labelling of acid-precipitable material was then determined. Changes in labelling due to variations of culture conditions were interpreted according to whether they were specific for control or antigen-treated cultures or non-specific. Cell concentration and serum concentration were predominantly non-specific variables influencing the labelling in control and antigen-treated cultures to a proportionate extent. However, at serum concentrations below 5 per cent labelling was disproportionately inhibited in antigen-treated cultures; there were further minor disproportionate inhibitions at higher serum concentrations. Labelling was inhibited by increasing the concentration of serum from 25 to 50 per cent, mainly due to a non-diffusible competitive inhibitory activity. Isotope-dilution analysis of the effects of serum on labelling over a wide range of serum concentrations indicated that the relationship was a complex one with at least three step-wise stimulations of the maximum labelling rate being produced by increasing the serum concentrations from 0 to 25 per cent. Labelling in antigentreated cultures containing post-immunization serum was less than labelling in cultures containing an equal volume of preimmunization serum, but labelling in control cultures was enhanced by post-immunization serum. These results are shown to be compatible with the proposals (i) that labelling in control cultures reflects the response of cells to low concentrations of endogenous antigens, and (ii) that preimmunization serum and post-immunization serum contain ‘natural’ and ‘acquired’ antibodies respectively, which normally buffer cell-borne receptor sites against reaction with endogenous and exogenous antigens. [ABSTRACT FROM AUTHOR]

In order to investigate the role of lymphocyte function-associated antigen 1 (LFA-1) in lymphocyte adhesion to endothelial cells (EC), we have studied the adhesion of a LFA-1-deficient lymphoblastoid cell line, ICH-KM, which has < 10% of the cell surface LFA-1 expressed on a normal lymphoblastoid cell line, ICH-BJ. The adhesion of ICH-KM cells to unstimulated EC was 49.9 ± 8-6% (mean ± SD) that of ICH-BJ cells. Moreover, phorbol ester-stimulated ICH-KM cells showed a considerably weaker increase in adhesion to unstimulated EC compared with ICH-BJ cells (mean ± SD increase in percentage adhesion, 3.8±2-3 compared with 18.5±8.0; P<0.025). In contrast, there was no significant difference between the enhanced adhesion of ICH-KM cells and ICH-BJ cells to interleukin-1 (IL-1)-stimulated EC. Thus ICH-KM cells showed a 22.7 ± 11.0 (mean ± SD) increase in percentage adhesion to IL-1-stimulated EC compared with the 24.8 ± 8-5 increase in percentage adhesion of ICH-BJ cells. Anti-LFA-1 monoclonal antibodies had no effect on the enhanced adhesion of ICH-KM and ICH-BJ cells to IL-1-stimulated EC but abolished the differences in adhesion between the two cell lines. The study therefore indicates that although a major part of unstimulated and phorbol ester-stimulated lymphocyte-EC adhesion is dependent upon LFA-1, the enhanced adhesion due to stimulation of EC with IL-1 is not dependent upon this molecule. The data therefore supports the existence of cytokine-inducible LFA-1-independent adhesion molecules for lymphocytes on EC. [ABSTRACT FROM AUTHOR]

The helper function of human T4+ cells acting on autologous B cells, in the presence of monocytce-enriched cells (MEC), has been studied using the hapten--carrier conjugate DNP--streptococcal antigen (DNP-SA). The antibody response can be suppressed by T8+ cells, but selection of a Vicia villosa lectin-non-adherent (T8VV-) subset enhances the suppresor function. The T8 Vicia villosa lectin-adherent (T8VV+) cells are not suppressive, rather they inhibit the T8VV- cell suppressor function. Sequential reconstitution studies suggest that the target of both T8VV- suppressor-cell and T8VV+ contrasuppressor activity is the T4 helper-inducer cell. This was established after the separation of T4+ cells into Leu 8- helper- and Leu 8+ suppressor-inducer cells. Activation of T8VV- and T8VV+ cells with antigen before reconstitution with fresh lymphocytes and MEC suggests that T8VV- suppressor activity is antigen specific, whereas T8VV+ cell contrasuppressor function is non-specific. We suggest that the human T8+ cells are functionally heterogeneous and consist not only of suppressor and cytotoxic cells but also contrasuppressor cells. [ABSTRACT FROM AUTHOR]

The role of suppressor cells in control of persistent infections may be of profound importance. Whether a positive immune response or suppression of immunity is generated at the time of initial exposure to pathogens causing such infections may in part be due to the nature of the initial antigen presentation to the specific cells of the immune system. We have shown that in vivo modulation of epidermal APCs by an environmentally encountered stimulus (UV-B light exposure) and subsequent transfer of these APCs together with live HSV-1 to naive syngeneic recipients is sufficient to generate suppression of DH rather than DH to HSV-1. This suppression is T-cell mediated and specific for HSV-l. The phenotype of the Ts cell induced by epidermal cell transfer is Thyl+ L3T4+ Ly2-. [ABSTRACT FROM AUTHOR]

The role of L3T4 antigens in the concanavalin A (Con A) response of regenerating spleen cells (Cy-SCs) after cyclophosphamide (Cy) treatment was studied. Anti-L3T4 monoclonal antibodies (Mabs) markedly inhibited the Con A response of the regenerating Cy-SCs, which do not require Ia molecules expressed on accessory cells (ACs) for Con A activation. However, the Con A response of normal spleen cells (N-SCs), which do require Ia molecules on ACs, was not inhibited by the same Mabs, although the Con A response of N-SCs, as well as that of Cy-SCs, was demonstrated to be mediated by L3T4+ T cells. The optimal times for the inhibitory effect of anti-L3T4 Mab was 7 days after Cy treatment, when the number of spleen cells increased to a maximum following a regenerative phase. Its inhibitory effect was reduced by high concentrations of Con A, and was restricted to the early phase of the Con A response. A short time exposure of the Cy-SCs to the anti-L3T4 Mabs was sufficient to decrease the response to Con A. Our results cannot explain the hypothesis that the L3T4 molecule functions solely by interacting with non-polymorphic parts of Ia molecules on ACs Taken together, these results and those of other groups of investigators suggest that Con A-induced T-cell activation may be mediated by at least two or more interaction mechanisms involving either Ia or L3T4 molecules. Firstly, normal L3T4+ T cells may mainly interact with Con A involving self Ia molecules on the ACs. The extent of this interaction is sufficient to induce T-cell activation, and then does not need another L3T4 molecule. Secondly, the regenerating L3T4+ T cells may usually internet with the cell surface antigens of other T cells, including L3T4, by the binding of both cell surface molecules to Con A in the absence of ACs, and then transmit a signal for 1-cell activation. Anti-L3T4 Mabs may exert inhibitory effects somewhere in this process. [ABSTRACT FROM AUTHOR]

Affinity maturation was studied by the analysis of the kinetics of the appearance of antibody subpopulations with different affinities during the immune response, using an hapten-inhibition ELISA. The immune response in KLH-Ar-immunized A/J mice was used as a model system. Five antibody subpopulations of different affinity (103-107 M-1) could be detected, the relative concentrations of which changed during affinity maturation. The high-affinity antibody subpopulations did not represent the major fraction at any stage during affinity maturation. The appearance of the highest affinity subpopulation (107 M-1), despite exhibiting relative concentrations no higher than 12%, produced an important increase in average affinity. On the other hand, its disappearance at the end of the maturation process could explain the average affinity decrease observed at this stage. Our results indicate that affinity maturation cannot be explained by the dominance of high-affinity clones, as proposed by Siskind & Benacerraf (1969). The increase in affinity could rather be due to the progressive appearance of low percentages of high-affinity clones, which are not present in the primary response and never become dominant. [ABSTRACT FROM AUTHOR]

The idiotypic profile of anti-phosphorylcholine plaque-forming cell responses and their evolution with ageing were studied in (NZB × NZW) F1 mice. Our results showed that the anti-phosphorylcholine plaque-forming cell response induced by phosphorylcholine coupled to keyhole limpet haemocyanin and, paralleling, the T15 idiotype clonal dominance declined with ageing. This loss of immune competence was also observed with another thymus-dependant (phosphorylcholine coupled to egg globulin) as well as thymus-independent (capsular polysaccharide of Streptococcus pneumoniae strain R36a) antigens. In contrast, old mice challenged with an antigenic preparation of Neisseria meningitidis showed an immune response not significantly different from that elicited by the same antigen in young mice. The hapten-augmentable plaque-forming cells were assayed to determine whether a putative auto-antiidiotypic regulation underlies this loss of immune competence. Only minimal numbers and non-significant differences between young and old mice immunized with any antigen could be detected. Further studies using an adoptive transfer system demonstrated that cells from aged mice were able to support a normal anti-phosphorylcholine response when transferred into lethally irradiated young recipients. Our results suggest that no permanent cellular defects, but rather internal environment or/and radioresistant suppressor cells, are involved in this loss of immune competence. The role played by these factors and their effect on distinct subpopulations of B cells are discussed. [ABSTRACT FROM AUTHOR]

The response of primed T cells to keyhole limpet haemocyanin (KLH) was used to compare the characteristics of antigen presentation by lymphoid dendritic cells, splenic and peritoneal macrophages. In a similar manner to macrophages, purified dendritic cells could be pulsed with antigen and subsequently fixed by brief glutaraldehyde fixation and still retain antigen presenting activity. Also, as previously reported for macrophages, presentation could be inhibited by chloroquine. These functional experiments suggested that the pathway of antigen presentation in dendritic cells and macrophages was similar or identical. However, biochemical studies, using radiolabelled antigen, showed that dendritic cells do not significantly degrade large proteins such as KLH to TCA-soluble form, but partially hydrolyse them to smaller peptide fragments. The significance of these results in terms of a model of the cellular pathways of antigen presentation is discussed. [ABSTRACT FROM AUTHOR]

Incorporation of lipid A into liposomes markedly enhances the plaque-forming cell (PFC responses against model membranes that are sensitized with synthetic amphipathic antigens 2,4-dinitrophenyl-6-N-aminocaproyldipalmytoylphosphatidylethanolamine (DNP-Cap-DPPE) and fluorescein thiocarbamyldipalmytoylphosphatidylethanolamine (FI-DPPE). The enhancing effect was pronounced only when lipid A and haptenic determinants were presented on the same liposome. Lipid A was incorporated into heterologous erythrocytes and syngeneic tumour cells. Significant and selective increase of in-vitro PFC responses was observed against heterologous erythrocytes, and immunization of C578L/6 mice with lipid A treated X-irradiated tumour cells (EL-4) markedly prolonged the survival of mice after challenge with homologous viable tumour cells. These data suggest that passively incorporated lipid A will be an effective method for selectively augmenting antibody formation against membrane-associated antigens. [ABSTRACT FROM AUTHOR]

The influence of antibody isotype on the lifetime of complexes involving cell surface antigens of the rat fibrosarcoma HSN.TC has been investigated using direct binding and competitive RIAs to monitor the antibodies. Alloantibodies of the lgG class that had bound to the cells during a 1-hr exposure to antiserum were cleared subsequently from the cell surface by an active process involving two distinct phases. Between 30 and 70% of these antibodies were lost in the first 10 hr but the antibodies remaining were cleared more slowly with half-lives ranging from 20 to 40 hr. Antibodies of the 1gM class, however, and those that could bind Clq and initiate the complement cascade were cleared rapidly with half-lives of less than 3 hr. Analysis of total cell-associated immunoglobulin showed that the disappearance from the cell surface was not a consequence of intracellular accumulation of antibody but was caused by the release of the antibody in a degraded form. The surface expression of the majority of the alloantigens involved was not affected by continuous exposure to antibody although modulation of a subpopulation of antigens could not be excluded. These results suggest that the clearance of alloantibodies involved their internalization and degradation and that antibodies capable of forming multimeric complexes were cleared rapidly. Some of the lgG-containing complexes, however, exhibited extended lifetimes at the cell surface, suggesting that either they were not internalized or they were recycled between the cell interior and the plasma membrane. [ABSTRACT FROM AUTHOR]

The effect of pregnancy on the immune status of CBA mice has been measured in terms of Ig-secreting cells. Reversed plaque assays show that there is a considerable increase in numbers of IgG- and, to a lesser extent, IgM-secreting cells in the para-aortic lymph nodes. Similar but muted response were observed in the spleen and among the lymphocytes of the peripheral blood. Analysis of cells sorted for intensity of staining for surface Ig in a fluorescence-activated cell sorter (FACS), shows that Ig-secreting cells are confined to a population of very weakly staining cells. The 'response' in syngeneic pregnancies (CBA × CBA) is at least as great as that seen in allogeneic pregnancies (CBA × C57). Active immunity to a xenogeneic erythrocyte antigen is potentiated by pregnancy. [ABSTRACT FROM AUTHOR]

A study has been made of the variables concerned in allotype suppression of adult spleen cells in the adoptive transfer system. These are: SRBC (antigen) dose; the dose and timing of injection of anti-allotype serum IgG; the number of spleen cells transferred and whether these cells were taken from primed or unprimed donors. Adoptively transferred primed cells are considerably less susceptible to suppression by concomitantly injected anti-allotype serum IgG than are unprimed spleen cells. Injection of anti-allotype serum during the period after adoptive transfer, has shown that primed cells loose their susceptibility sooner (2 days) than the unprimed cells (4 days). Allotype heterozygous CBA spleen cells arc less susceptible to allotype suppression than either allotypically homozygous or heterozygous non-H-2k cells (H-2b,d, or s). Allotype suppression of the TI IgG response to DNP-Ficoll was measured 7 days after adoptive transfer of allotype-homozygous cells from both normal and nude CBA mice (unprimed). The results indicate that T cells do not play a role in the initiation of short-term allotype suppression in the adoptive transfer system. [ABSTRACT FROM AUTHOR]

Lymphocytes reactive against cultured cerebellar cells were collected from rats that had been sensitized against a variety of preparations. While cells with in vitro reactivity were invariably present in lymph from rats which were to develop encephalomyelitis, the reverse did not apply. Challenge with either neural or non-neural allogeneic tissue sensitized effectively against allogeneic neural tissue. Challenge of homograft-tolerant rats with either syngeneic or tolerated allogeneic spinal cord sensitized against neural tissues of tolerated strain. [ABSTRACT FROM AUTHOR]

Following previous authors, the term antigenic modulation is used to describe the induction, by antibody, of resistance to lysis by antibody plus complement. A report is given of the rapid antigenic modulation in vitro of surface immunoglobulin (Ig) on guinea-pig L2C leukaemic lymphocytes: incubation of the cells for 2 min or longer at 37° with anti-Ig diminished or removed completely the lysis occurring during subsequent incubation with anti-Ig plus complement. The modulation was effective for both xenogeneic (rabbit) and syngeneic (guinea-pig strain 2) complements, but more rapid for the latter. It appeared simply to require the action of antibody on a metabolically active cell: no requirement could be demonstrated for any serum component other than antibody, and there was a need to raise the temperature to 37° after attachment of the antibody. There was molecular specificity inasmuch as modulation with anti-Ig failed to confer any resistance to lysis by another antibody (anti-Ia) plus complement. [ABSTRACT FROM AUTHOR]

Endotoxin (1–10 μg intravenously) increased the number of strongly peroxidase-positive cells in the white pulp of the mouse spleen, demonstrable by the Graham Karnovsky technique. The increase was greatest after 10 h, the cells being found most frequently around the central vessels of the white pulp. The appearance of such cells at this critical immunological site may relate to the known adjuvant activity of endotoxin in promoting immune responses against protein antigens. [ABSTRACT FROM AUTHOR]

Passage of spleen or bone marrow cells from non-immunized donors through a DNP-Ova coated Degalan bead immunoadsorbent column resulted in a specific depletion of the ability of the cell population to restore the anti-DNP antibody response to DNP-BGG by lethally irradiated mice. There was a preferential depletion of precursors of high affinity antibody-secreting cells which is consistent with the view that these cells bear high affinity antigen receptors The cell population passed through the immunoadsorbent column was also preferentially depleted of the capacity to produce high affinity direct anti-DNP PFC in comparison with high affinity indirect anti-DNP PFC These findings suggest, but do not prove, that different subpopulations of antigen. binding cells are precursors for direct and indirect PFC. [ABSTRACT FROM AUTHOR]

The intensity of the cellular immune response to strong transplantation antigens of the H-2 complex is related to the histological type of cells taken for immunization. Splenic cells from C3H mice immunized BALB/c mice better than cells of non-lymphoid origin from the same donor. The reverse was observed when, instead of BALB/c mice, C57B1/6 and A/He were used in immunization experiments. Analysis of the inheritance of this immunization pattern studied in mice of the first generation from various crosses between strains (BALB/c, C57B1/6 and A/He) and in mice from backcrosses has demonstrated that this immunization pattern is not inherited as a simple Mendelian character. Therefore, it could be supposed that efficiency of immunological control of cells having different histological types, may be genetically determined. [ABSTRACT FROM AUTHOR]

The delayed-type-hypersensitivity (DTH) reaction observed in mice primed i.v. with low doses of sheep red blood cells is not detectable after priming with large doses. Estimation of the number of DTH mediating cells (DTH-C) was performed by titration (D50 determination) using local adoptive transfer of these cells mixed with antigen into footpads of unprimed mice. In opposition to the absence of peripheral DTH-C, splenic DTH-C were more numerous after large doses than after low doses of antigen. The ability of cyclophosphamide lo restore DTH responsiveness in mice primed with large doses appeared to be related to an increased cirulation of DTH-C. These cells circulated only in mice without any detectable B-cell response or in which this response was depressed. Thus a B-cell dependent splenic retention process of DTH-C is discussed as the origin of DTH unresponsiveness observed after priming with large doses of antigen. [ABSTRACT FROM AUTHOR]

We have investigated the ability of male-mouse spleen cells before and after culture in the absence of deliberate antigenic stimulation to show specific cytotoxicity to syngeneic embryo-fibroblast cells. The data suggest that cytotoxicity which develops spontaneously in such spleen cell cultures is directed primarily against embryo-associated antigens. Syngeneic Con-A-stimulated spleen cells, which, unlike fresh normal spleen cells, are also lysed by rabbit anti-mouse embryo antisera, are also a suitable target to demonstrate spontaneously developing cytotoxicity. [ABSTRACT FROM AUTHOR]

A fowl antiserum to human polyclonal F(ab')2 with specificities for variable region antigens of both light and heavy chains and for the constant portion of the Fd fragment bound to about 20 per cent of thymocytes from three children. Binding was completely inhibited by purified human poly- I clonal IgG and only partly by L chains. The fowl anti-F(ab')2 also bound to a subpopulation of peripheral blood lymphocytes which was somewhat larger than the B-cell population identified with other anti-immunoglobulin sera. Binding was detected by autoradiography and the amount of radiolabel seen on thymocytes was nearly as great as that on peripheral blood lymphocytes. As no binding to thymocytes was seen with other anti-immunoglobulin sera we propose that the fowl I anti-F(ab')2 recognizes immunoglobulin or material with structural similarities which is synthesized by thymocytes but of which predominantly V region determinants are exposed on the membrane surface. [ABSTRACT FROM AUTHOR]

The gastric parietal cell auto-antigen is localized essentially in the microsomal fraction of mucosal homogenates obtained by centrifugation over a sucrose density gradient. The fractionation is facilitated by digestion of gastric mucus with ficin before disruption of the tissue. The antigen was assayed by a quantitative complement-fixation method. The effect of a number of enzymes and chemical treatments upon the serological reactivity of a gastric microsomal preparation was studied. The properties of the gastric antigen suggest an intimate association with insoluble lipoprotein elements of the microsomal membranes. Although immunologically distinct and strictly organ specific, the cytoplasmic auto-antigens of the stomach, and of the thyroid gland, show a remarkable resemblance in their biochemical characteristics. This similarity has a parallel not only in the embryological origin and the iodide trapping mechanisms which the two organs share in common but also in the close serological and clinical associations that have been demonstrated between auto-immune atrophic gastritis and thyroiditis. [ABSTRACT FROM AUTHOR]

Rabbits were immunized with bovine (BSA) or human (HSA) serum Albumin. Spleen cell suspensions were prepared and incubated with the homologous or cross-reacting albumins. The antigen-dependent stimulation of the rate of DNA synthesis was used as a measure of the antigenic activity of each of the albumin preparations. The response was proportional to the logarithm of the antigen concentration for both the homologous and the cross-reacting albumins and the slopes of each curve were approximately the same. The cross-reacting albumins were less stimulatory than the homologous antigen. The pattern of their relative activities in this system tbllowed the same pattern shown in their serological cross-reactions. [ABSTRACT FROM AUTHOR]

Rabbits injected with a homogenate of rat gut mucosa produce auto-antibodies reacting with their own gut mucosa. Different organ-specific antigens present in the stomach mucosa and colonic and ileal mucosae can be demonstrated in this way. The antigens concerned are in the mucous secretions as well as in certain epithelial cells and may be mucopolysaccharides. [ABSTRACT FROM AUTHOR]

Previous findings of the appearance of pyrogenic activity when spleen and lymph node cells from BCG-sensitized rabbits were incubated with tuberculin in vitro have been repeated and extended in experiments with guinea pigs. Two different systems were used: hypersensitivity induced by BCG infection and by means of diphtheria toxoid-antitoxin precipitates. Cells were incubated in vitro with diluted tuberculin, diphtheria toxoid or Hanks's solution. Supernatant fluids obtained after various incubation periods were tested in normal guinea pigs for their pyrogenic activity and for ability to induce skin inflammatory reactions. Pyrogenic substances were obtained in most of the experiments in which cells of hypersensitive animals were incubated with the specific antigen; experiments with the unrelated antigen and controls were negative. Formation of skin-irritating substances was less regular, and they were sometimes formed non-specifically in control experiments also; in general, however, the results of these experiments agreed with the results of the fever reaction. The results obtained are discussed in relation to the mechanism of delayed hypersensitivity. [ABSTRACT FROM AUTHOR]

These studies have tested the behaviour of antibody-producing cells on serial transfer through immunologically neutral hosts. Spleen cells from rats primarily immunized in vivo with a Salmonella flagellar antigen were secondarily stimulated in vitro by incubation with the antigen at 37° C. They were then injected intraperitoneally into homologous rats less than 48 hours old. Two weeks later these host rats were killed, their sera titrated for antiflagellar antibody, and their spleen cells transferred to further newborn hosts, either with or without further antigenic stimulation. This type of transfer was continued for up to seven passages. It was found that if the spleen cells were not antigenically stimulated in vitro, they formed comparatively small quantities of antibody, and successful passage could only be achieved for two or three passages. If the spleen cells were antigenically stimulated before passage, much more antibody was produced, and successful passage could be achieved for five passages. Moreover, if the donor animal had been immunized with two unrelated antigens, and the spleen cells before passage were stimulated with one antigen but not the other, the hosts developed far more antibody to the former than to the latter. These results are discussed in the light of recent immunological theory, and are considered to support clonal hypotheses of antibody formation. [ABSTRACT FROM AUTHOR]

Sheep red cells (RC) were coupled with HSA by means of an HSA-anti-SRBC IgG conjugate. The coupled RC were used to prepare rosettes with spleen cells of mice immunized to HSA in Freund's complete adjuvant. The rosettes were fractionated on discontinuous Ficoll gradients and the rosette-rich and rosette-poor fractions were tested for ability to transfer adoptively secondary responsiveness to HSA. Efficiency of transfer was measured in terms of haemagglutination titres. Memory of HSA was shown to be associated with rosette-forming cells, i.e. with cells carrying receptors for HSA. Several parameters of the adoptive transfer system employed were studied. The HSA-coupled RC proved to be very efficient in stimulating secondary responses to HSA. High titres of haemagglutinating antibodies (up to 1/80,000) were obtained with 10 × 106 spleen cells transferred and 2 × 106 spleen cells consistently gave titres of the order of 1/10,000. [ABSTRACT FROM AUTHOR]

The induction and loss of tolerance to native and depolymerized fractions of levan, a thymus-independent antigen, have been studied in (DBA/1 × CBA-T6T6)F1 mice by a direct plaque-forming cell (PFC) assay. Tolerance following 1 mg native levan began to break by 80 days, whereas mice remained specifically unresponsive 150 days after 10 mg. Effects attributable to long persistence of antigen have been demonstrated. The following main features of levan tolerance were established by transfer of spleen cells to 900 R-irradiated recipients: (1) the induction period is very short—less than 24 hours. (2) No detectable immunity accompanies or precedes induction. (3) The fall in immunogenicity with reduction in molecular weight (mol. wt) of levan is not accompanied by loss of tolerogenicity. An effectively tolerogenic, non-immunogenic fraction (mol. wt< 10,000) was found. (4) There is no rapid loss of tolerance following cell transfer (reactivity is only regained after 50 days), arguing against the existence of tolerant cells in this model. As these results and those from corresponding investigations with SIII and HGG all show major dissimilarities, it is considered imprudent to base generalizations concerning the nature of B cell tolerance on studies with only one type of antigen. Whether or not two different kinds of specific unresponsiveness might be implicated with these cells is discussed. [ABSTRACT FROM AUTHOR]

The immunogenicity of antigens associated with peritoneal exudate cells was compared with the immunogenicity of free antigens for the induction of delayed hypersensitivity. Oil induced peritoneal exudate cells were incubated with antigens labelled with radioactive iodine, washed and injected intraperitoneally into recipient guinea-pigs. Comparable amounts of free antigens were also injected. All sixteen inbred guinea-pigs given human or bovine gamma-globulin associated with peritoneal exudate cells gave positive reactions (greater than 5 mm) while the eighteen guinea-pigs given free antigens failed to show a reaction. Similar results were obtained with serum albumins in inbred guinea-pigs. The increased immunogenicity of antigens associated with peritoneal exudate cells was also seen in outbred guinea-pigs. Pretreatment with Freund's complete adjuvant did not depress the immunogenicity of antigens associated with peritoneal exudate cells and actually increased the induration of the 24-hour reaction to picrylated human scrum albumin and human serum albumin associated with peritoneal exudate cells. [ABSTRACT FROM AUTHOR]

Investigations of surface antigens of animal cells require sensitive and quantitative in vitro methods. A modification of the mixed haemadsorption technique (MHT) suitable for such purposes is presented. The indicator cells have been labelled with chromium-51. During studies of experimental conditions, including reproducibility and sensitivity, the technique was used in four model systems including histocompatibility (H-2, HL-A), species, organ and blood group antigens. Isotope labelling of the indicator cells has rendered the test more sensitive and in addition some qualitative properties of antigen-antibody interaction have been studied. The test procedure is simple and fast. [ABSTRACT FROM AUTHOR]

The effect of prior administration of two classes of 7S mouse anti-sheep red blood cell (SRBC) antibodies on the primary immune response to SRBC was studied. Mouse γG1 and γG2 immunoglobulins were isolated from serum by zone electrophoresis and density gradient isolectric focusing. The immunoglobulins were defined by qualitative and quantitative studies, and their effects on the primary response were determined by administering doses ranging from 0.5 to 0.001 mg of immunoglobulin 2 hours before injection of antigen. Gamma G1, antibody suppressed the response to SRBC at all concentrations. High doses of γG2 antibody partially suppressed 19S plaque-forming cells (PFC), but had no significant effect on serum haemagglutinin (HA) levels. Low doses of γG2 antibody specifically augmented both the 19S PFC and serum HA levels. It is concluded that the specific suppressing and augmenting influences of antibodies on the primary response are a function of class. In addition, it is proposed that &gammaG1, and γG2 antibodies can aet as specific regulatory elements during the primary response by exerting these two competing biological effects on antibody synthesis. [ABSTRACT FROM AUTHOR]

The primary antibody response in rabbits to human serum albumin (HSA) has been analysed. Doses of 0.04-25 mg were given subcutaneously with Freund's complete adjuvant. Maximum antibody response about 60 days after primary stimulation was independent of the antigen dose given. After 175 days the amount of persisting antibody was lowest in the animals stimulated with the lowest dose of antigen. The avidity of the antibody was inversely proportional to the antibody titre. Secondary stimulation of all rabbits with 1 mg of HSA intravenously caused secondary antibody responses which were insignificantly different from each other. It is suggested that the magnitude of the secondary antibody response is determined by the regulatory activity of the circulating antibodies following primary stimulation. [ABSTRACT FROM AUTHOR]

Both differentiating plasma cells and modified lymphocytes (lympho-plasmocytes) synthesize specific humoral antibody in rabbits, rats, mice and guinea-pigs. They do so during both primary and secondary responses, after subcutaneous injection of three enzymic antigens: horse radish peroxidase, glucose oxidase and alkaline phosphatase, administered as soluble or insolubilized proteins or alum precipitated or mixed with Freund's adjuvant. [ABSTRACT FROM AUTHOR]

Ficoll, a polymer of sucrose, has several advantages over proteins for preparation of density gradients used to study mammalian cells. In addition to its ease of preparation, Ficoll as a result of being an uncharged molecule, does not bind ions from solution and, therefore, does not affect the osmolality of the medium in which it is dissolved. In studies on antibody-producing cells we found that the measurement of buoyant density in Ficoll at pH 7.2 gives an accurate measurement of cell density. In such gradients antibody-producing cells band in a single peak at a density of 1.070 g/cm³. Two factors have a profound influence on the density profile of antibody-producing cells. Lowering the pH to 5.5 lowers their buoyant density to 1.062 g/cm³. Creation of an inverse osmotic gradient varying from 294 m-osmolal at the top of the gradient to 286 m-osmolal at the bottom splits the single peak of antibody-producing cells into three distinct peaks. This effect of osmotic gradients could account for the multiple peaks observed when cells are separated on protein gradients. [ABSTRACT FROM AUTHOR]

The mixed haemadsorption test was applied to cultured cells grown in Falcon plastic ‘Microtest’ plates. This small-scale method made it possible to simplify the whole procedure, to take many tests simultaneously, and in addition, to save both the amount of antisera and cells. Therefore, the method is suitable for titration of allo-antibodies against surface antigens and for simultaneous demonstration of the expression of different surface antigens. [ABSTRACT FROM AUTHOR]

A small scale modified method of the immune adherence on monolayer cells grown in Falcon plastic ‘Microtest’ plates is described. This method is especially useful in mouse systems as it simplifies the washing procedure which is an indispensable step for removal of the anticomplementary activity present in mouse serum. Histocompatibility antigens and the surface antigen induced by the Moloney virus could be detected with this method. [ABSTRACT FROM AUTHOR]

Summary. A comparison has been made between the reactivity of cross-reacting antibody from group O sera titrated against A cells and the reactivity when titrated against cells of group B. It is considered that the experimental findings require the postulation that cross-reacting antibodies which are produced in response to stimulation by antigen A have a different configuration from those produced in response to antigen B. The results also suggest that elution of antibody from cells at 56° may cause minor changes in spatial configuration which diminish the ability of some antibodies to combine with some subgroups of the A system. [ABSTRACT FROM AUTHOR]

Antisera to specific proteins in herpes simplex infected cells were produced by immunization of rabbits with infected rabbit kidney cells. These antisera were highly virus specific and produced up to twelve lines in immunodiffusion tests against infected cell extracts. Acrylamide electrophoresis and immunoelectrophoresis revealed up to ten virus specific proteins of varying size. [ABSTRACT FROM AUTHOR]

A synthetic multichain polypeptide, (T,G,)-A--L 509, was trace labelled with 125I in the tyrosine end groups, composing the main antigenic determinants, and used to study the distribution of antigen in the draining lymph nodes after injection into the footpads of mice. The polypeptide was administered: (a) in saline solution into unprimed mice—in which form it is not demonstrably immunogenie; (b) in Freund's adjuvant into unprimed mice—in which form it is immunogenie; and (c) in saline solution to primed mice, so as to give a booster response. Experiments comparable to (c) were also done with [125I]haemocyanin. At various time intervals from 12 hours to 21 days later sections of the draining nodes were examined by radioautography and methyl green—pyronine staining, or by a combination of immunofluorescent staining for antibody containing cells with radioautography. In unprimed mice, irrespective of whether they made antibody, labelling was found predominantly in tile phagocytic cells of the medulla and the cortical sinus, but definite weaker labelling was also seen in the germinal centres. The label persisted throughout the period of observation, but tended to become more prominent with time in the germinal centres of mice making antibody m response to antigen m Freund's adjuvant. In primed mice the label was rapidly and predominantly concentrated in germinal centres, where it persisted throughout the period of observation while its intensity gradually diminished in other sites. The distribution of label in germinal centres was in a lace-like or dendritic pattern, similar to that described by Nossal and co-workers, and did not correspond to the lymphocytes or pyroninophilic cells in the centres. Grain counts were made over specific antibody containing cells in both primary, and booster responses to (T,G)-A--L. Such cells sometimes lay close to and sometimes many cell diameters distant from phagocytic cells containing concentrations of the antigen, but the cells themselves did not have grain counts significantly different from the background count. If the antigen remained undegraded, under the conditions of the experiment it would have been possible to detect the presence of fifteen molecules or less in a cell. Reasons are given for supposing that 125I constituted a valid label of the main antigenic determinant groups, of which there were a maximum of 100 per molecule. [ABSTRACT FROM AUTHOR]